OncoLab Diagnostics Targets Cancer Recurrence Monitoring with CTC-Derived Biomarker Assay

NEW YORK – Austrian firm OncoLab Diagnostics is developing a range of liquid biopsy-based diagnostic products, starting with an assay for monitoring for late-stage cancer recurrence.

OncoLab expects to receive a CE mark later this year for its RT-qPCR test, which detects and characterizes epithelial CTCs that express cytokeratin-19 (CK-19) as a biomarker. In addition, the firm is leveraging a recently signed partnership with Axon Dx to improve assay development.

OncoLab was founded in 2010 by CEO and Chief Scientific Officer Robert Zeillinger to develop diagnostic tests for the oncology space. The company initially struggled to identify "the right partners and investors" to help fund its commercial plans, but in early 2019 it signed a contract with an undisclosed private investor and began commercializing its CTC detection technology.

While Zeillinger is a professor of tumor biology at the Medical University of Vienna, he noted that OncoLab developed its technology as a separate commercial entity and did not spin out from the university. 

Zeillinger explained that a user of OncoLab's test kit would collect 10 ml of a patient's blood sample, and remove red blood cells through a density centrifugation step. The kit then lyses the remaining cells and isolates their RNA, followed by RT-qPCR to identify and quantify the presence of CK-19 and other CTC-associated biomarkers.

"CK-19 is not expressed in white cells, [and thus] reduction of white blood cells is not as critical," Zeillinger said. "Most of the research comes from breast cancer, so [CK-19 was] an ideal marker for breast cancer CTC detection." 

Zeillinger said that OncoLab's RT-qPCR test can produce results within 24 hours of collecting a patient blood sample.  

Zeillinger declined to disclose information about the RT-qPCR assay's clinical sensitivity and specificity, but highlighted that multiple published research papers on the use of CK-19 for breast cancer detection indirectly validate the firm's approach.   

Max Wicha, professor of medicine and oncology at the University of Michigan's Rogel Cancer Center who is not involved with OncoLab, pointed out that CK-19 has been used as a biomarker for breast cancer detection for several years. Researchers have developed CK-19 assays and compared them to Menarini Silicon Biosystems' CellSearch CTC enumeration assay, the only CTC-based diagnostic test that has received 510(k) approval from the US Food and Drug Administration.

However, Wicha argued that CK-19's major limitation is that the protein is not a clear cancer biomarker, but instead a marker for circulating epithelial cells.

"If you look at the level of CK-19 in patients … even with patients with non-metastatic breast cancer, there's [about a] one- to two-fold increase in CK-19," Wicha said. "You even find [levels of CK-19] in normal women, because they shed [epithelial cells] in their blood."

Wicha believes that it is still unclear whether tools using CK-19 as a biomarker for detecting cancer recurrence "are stronger" than current detection methods like CellSearch.

"CK-19 tends to be positive when CellSearch is positive, but the researchers will need to show that when CK-19 is positive and CellSearch is negative, the patients will go on to develop metastatic cancer," Wicha said.

Zeillinger acknowledged that OncoLab has dealt with multiple logistical and technical challenges in developing a multi-biomarker version of the assay to characterize the CTCs.

"Some of these assays might test for somewhere between 5 to 50 biomarkers, but you can't include 50 in a RT-qPCR assay," Zeillinger said. "Because it's impossible to consider all of these biomarkers, we have to decide which markers to combine or ignore for our assays."

In addition, Zeillinger noted that when OncoLab wants to incorporate more than 12 biomarkers in a future version of the assay, the test's workflow will require pre-amplification steps. The firm has therefore experimented with methods such as Angle's Parsortix system and Vortex Bio's VTX-1 technology to enrich blood samples for the multi-marker assays.

Oncolab currently offers the breast cancer assay for research-use-only purposes in Europe and the US, and anticipates obtaining a CE mark and launching an in vitro diagnostic version later this year.

While the RUO assay is platform-agnostic, Zeillinger said the CE-IVD version, which he envisions will cost less than €150 ($182) and be called "DX4U", will run on Roche Diagnostics' LightCycler platform. The firm also plans to eventually apply for FDA 510(k) clearance for the assay. 

OncoLab will have to compete with multiple players in the space that are also developing or offer liquid biopsy methods to monitor for cancer recurrence. Wicha noted that the cancer diagnostic field is shifting toward the use of circulating tumor DNA (ctDNA)-based platforms, such as Guardant Health's ctDNA assay, to monitor for cancer recurrence. Wicha's own team at the University of Michigan is looking into using cancer cell-derived RNA to identify cellular pathways that are active during metastasis and can possibly be susceptible to targeted therapeutic agents. 

"[OncoLab] is not going to convince anyone to use the test unless they publish data that it's better than any of these other methods … and detect recurrence when both ctDNA and CellSearch are negative," Wicha said. "The future of these types of tests will involve capturing the cells and performing much more complicated analysis, not just whether the cells are there or not."

However, Zeillinger believes that OncoLab's platform stands out from other methods for cancer recurrence because it is the first to offer a CK-19-based RT-qPCR assay.

"CK-19 is probably the best marker we have for the detection of epithelial CTCs," Zeillinger said. "However, since several CK-19 pseudogenes exist, there were no good PCR tests for CK-19 for CTC analysis on the market, [but] our test changes this because our technology avoids the amplification of these pseudogenes."

Zeillinger expects that future versions of OncoLab's technology will use patented combinations of several RNA biomarkers to provide qualitative information about the patient's cancer. As part of the firm's pipeline, it will develop RT-qPCR assays to indirectly detect CTCs in ovarian, cervical, lung, prostate, and neuroendocrine tumors.

"We will have a special breast cancer test that provides more information about CTCs in breast cancer, which will give us more information about possible responses to therapy and improve the test's sensitivity," Zeillinger said.

However, Wicha argued that OncoLab will need to demonstrate through longitudinal studies that it can identify CK-19-positive patients who also progress and develop a metastatic tumor.  

"If you develop something that would predict how soon the patient will relapse, say a year before they would relapse and institute therapy, that would be valuable," Wicha said. "If you can show that very significant statistic earlier than other methods, then they might have something valuable, but again [OncoLab] will need to show the data."

OncoLab originally acquired a patent from the University of Greece for its CK-19-based product line, and now holds a patent family on the use of CK-19 expression in breast cancer and gynecology-based cancers, as well as other biomarkers for small-cell lung cancer and neuroendocrine cancer.

OncoLab has also partnered with Earlysville, Virginia-based Axon Dx to build immunofluorescent assays to identify and count CTCs. Zeillinger added that using technology individually patented by both firms, OncoLab and Axon Dx will develop the assays so they can help improve the prediction of patients' outcome and aid in selecting appropriate therapies.  

"We found out that if we combined the two methods together, then the correlation with the clinical data — regarding patient outcomes and their prognosis — significantly increased," Zeillinger said. "Therefore, to combine the two methods that allow detection and characterization of CTCs is a good thing."

Axon Dx CEO Kent Murphy noted that the firm first approached OncoLab in 2020 "to represent" its immunofluorescent technology in different markets. Axon Dx's research-use-only nCyte Dx platform scans all cells in a blood sample using an optical scanner, then uses artificial intelligence on the resulting images provide morphological and molecular characterization of rare cells such as CTCs, circulating endothelial cells, and circulating stromal cells.

The firms also signed a deal earlier this week under which OncoLab will distribute the nCyte Dx platform in the Eastern European, Middle Eastern, and African markets.

While Zeillinger believes Axon Dx's methods complements his team's RT-qPCR tests, he said that the firm will continue to experiment with additional CTC enrichment platforms in the future.